Abstract Biological AI learns primarily from existing observations, but many questions cannot be answered from available data alone. Here we show that AI can instead acquire knowledge by acting directly on biological systems and learning from the consequences. We developed a closed-loop framework in which autonomous agents design protein variants, construct and characterize them in a robotic laboratory, learn from the resulting experimental feedback, and decide what experiments to perform next. We then allowed the system to operate continuously and without human intervention for approximately one month, during which multiple agents independently explored protein sequence space while learning from shared experimental experience. Applied to glycoside hydrolases, the agents discovered enzymes with substantially altered substrate specificity toward non-native sugars and progressively learned the structure of the underlying sequence–function landscape. The resulting experimental experience also revealed determinants of substrate specificity and protein expression that were not specified as learning objectives. These results demonstrate that AI can autonomously interact with biology over extended periods to acquire knowledge through experience, establishing a framework for biological discovery driven by continuous experimental interaction.
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